Tackling Treg mediated resistance to radiation and anti-PDL1 in HNSCCs

解决 HNSCC 中 Treg 介导的放射抗性和抗 PDL1

• 批准号: 10590767
• 负责人: SANA KARAM
• 金额: $ 43.91万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10590767
• 关键词: Adjuvant Radiotherapy; Affect; Animal Model; Back; Biological Assay; Blood; Blood specimen; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CD8B1 gene; CXC chemokine receptor 3; CXCL9 gene; CXCR3 gene; Cell secretion; Cells; Chemotaxis; Clinical Trials Design; Coculture Techniques; Cytometry; Data; FOXP3 gene; Feeds; Flow Cytometry; Genetic Engineering; Genetic Transcription; Genetically Engineered Mouse; Goals; Growth; Growth Factor; Head and Neck Cancer; Head and Neck Squamous Cell Carcinoma; Human; Immune; Immune Evasion; Immune Targeting; Immune checkpoint inhibitor; Immunofluorescence Immunologic; Immunotherapy; In Vitro; Infiltration; Interferon Type II; Investigational Therapies; Knockout Mice; Knowledge; Laboratories; LoxP-flanked allele; Mediating; Modeling; Molecular; Mus; Neoadjuvant Therapy; PD-L1 blockade; Pathway interactions; Patients; Phase; Phase I Clinical Trials; Phenotype; Phosphorylation; Plasma; Proliferating; Radiation; Radiation therapy; Regulatory T-Lymphocyte; Relapse; Research; Research Proposals; Resistance; Resistance development; STAT1 gene; STAT3 gene; Signal Transduction; Signaling Molecule; T-Cell Activation; T-Cell Depletion; TCR Activation; Testing; Tissue Sample; Tissues; Up-Regulation; anti-CTLA4; anti-PD-L1; anti-PD-L1 therapy; anti-tumor immune response; cancer cell; carcinogenesis; checkpoint therapy; chemokine; clinical trial analysis; cofactor; cytokine; effective therapy; effector T cell; first-in-human; genome sequencing; high risk; immune checkpoint blockade; improved; inhibitor; inhibitor therapy; knock-down; migration; mouse model; negative affect; novel therapeutic intervention; pharmacologic; phase I trial; polarized cell; potential biomarker; programmed cell death ligand 1; recruit; response; synergism; targeted treatment; therapy development; therapy resistant; transcriptome sequencing; tumor; tumor eradication; tumor microenvironment; whole genome

Abstract Current treatment of head and neck squamous cell carcinoma (HNSCC) with immune checkpoint inhibitors and radiotherapy (RT) are initially effective, but relapse is common. The goal of our research is to understand mechanisms of response and resistance and to use this knowledge to improve therapy. We show that during the response phase to treatment, RT increases STAT1 phosphorylation, CXCL9/10 secretion, and PDL1 expression on cancer cells, and increases CD8 and CD4 infiltration and activation. These effects, however, dissipate in the resistance phase, and a significant increase in regulatory T cells (Tregs), STAT3, and TGF1 are observed. Treg depletion and anti-STAT3 targeting restores response to RT and anti-PDL1, resulting in suppression of plasma levels of TGF1, activation of T effector cells (Teff, Cd44+, IFN-G+ CD4 and CD8 T cells) and complete tumor eradication in an orthotopic model of HNSCC. This effect is unachievable with the addition of anti-CTLA4, anti-Tim3, or with hypofractionation of RT. Our in vitro preliminary data show that RT enhances STAT3 phosphorylation on CD4 T cells, leading to increased Treg conversion. This is reversed with STAT3 inhibitors. CD4 T cell conversion toward a Treg phenotype represents an important mechanism of immune evasion. TGF1 has been established as a necessary growth factor for conversion of naïve CD4 T cells to Tregs through induction of FoxP3 expression. In addition, STAT3 is a necessary transcription co-factor for FoxP3 expression. Our central hypothesis is that during the response phase, RT induces a distinct cancer cell STAT1-mediated CXCL9/10 secretion which recruits and activates Teff resulting in enhanced synergy with anti-PDL1. RT, however, also promotes CD4 conversion to Treg cells by phosphorylation of STAT3, which leads to the development of treatment resistance by inhibiting Teff function. Using knockdown constructs of STAT1 and CXCL9/10 on cancer cells and a CXCR3-/- mouse model, we will determine the effect of RT-induced chemokines in Teff chemotaxis, proliferation, and TCR activation (Aim1). To determine how RT affects CD4 T cell conversion and how that feeds back to negatively affect Teff function, we will use the CD4-Cre/ Stat3flox/flox mice with knockdown of STAT3 on CD4 T cells as well as STAT3 inhibitors (Aim2). Finally, we will use tissue and blood samples from a Phase I clinical trial aimed at using neoadjuvant combination RT and anti-PDL1 and will examine whether that changes the immune landscape to increase Teff/Treg ratio and enhance Teff activation profile (Aim3). We expect these studies to elucidate molecular and cellular parameters of RT plus immune checkpoint blockade and to help develop more effective therapy for HNSCC.

摘要 目前头颈部鳞状细胞癌(HNSCC)的免疫检查点抑制剂和 放射治疗(RT)最初是有效的，但复发很常见。我们研究的目标是了解 反应和耐药性的机制，并利用这一知识来改进治疗。我们展示了这一点 在治疗的应答期，RT增加STAT1的磷酸化，CXCL9/10的分泌和PDL1 在癌细胞上的表达，并增加CD8和CD4的渗透和激活。然而，这些影响， 在抵抗期消散，调节性T细胞(Treg)、STAT3和转化生长因子1显著增加 都被观察到。Treg耗竭和抗STAT3靶向恢复对RT和抗PDL1的应答，导致 抑制血浆转化生长因子1水平，激活T效应细胞(T细胞、CD44+、干扰素-G+CD4和CD8T) 细胞)，并在HNSCC的原位模型中完全根除肿瘤。这一效果是无法通过 加入抗CTLA4、抗TIM3或低分馏RT。我们的体外初步数据显示，RT 增强CD4T细胞上STAT3的磷酸化，导致Treg转化增加。这与以下情况相反 STAT3抑制剂。CD4T细胞向Treg表型的转化代表了 免疫逃避。转化生长因子1已被确定为转化幼稚CD4T细胞所必需的生长因子 通过诱导FoxP3的表达将细胞转化为Tregs。此外，STAT3是一种必需的转录辅助因子 用于FoxP3表达。我们的中心假设是，在反应阶段，RT诱导了不同的 癌细胞STAT1介导的CXCL9/10分泌，募集并激活TEFF，导致 增强了与抗PDL1的协同作用。然而，RT也通过以下方式促进CD4向Treg细胞的转化 STAT3的磷酸化，通过抑制Tef而导致抗药性的发展 功能。在癌细胞和CXCR3-/-小鼠上使用STAT1和CXCL9/10的敲除结构 模型中，我们将确定RT诱导的趋化因子在TEF趋化、增殖和TCR中的作用 激活(Aim1)。确定RT如何影响CD4T细胞转换，以及这种影响如何反馈为负 影响Tef功能，我们还将使用STAT3基因敲除了CD4T细胞的CD4-Cre/Stat3flx/FLOX小鼠 作为STAT3抑制剂(AIM2)。最后，我们将使用一期临床试验的组织和血液样本，旨在 使用新佐剂联合RT和抗PDL1，并将检查这是否会改变免疫 景观，以增加TJeff/Treg比率和增强Tef激活轮廓(Aim3)。我们预计这些研究将 阐明RT加免疫检查点阻断的分子和细胞参数，并帮助开发更多 HNSCC的有效治疗。